PROJECT SUMMARY ? PROTEOMICS AND METABOLOMICS SHARED RESOURCE The primary goal of the Proteomics and Metabolomics Shared Resource (PMSR) is to provide comprehensive proteomics and metabolomics services to Cancer Center members as well as other regional NCI-designated Cancer Centers (Fox Chase Cancer Center, Thomas Jefferson University Cancer Center), and secondarily to investigators in other academic institutions. Prior to December 2017, major mass spectrometry (MS) instruments in the PMSR included a Thermo Q Exactive HF, a Thermo Q Exactive Plus, and a Sciex QTRAP 5500 hybrid triple quadrupole mass spectrometer. These instruments have nano-UHPLCs (ultra-high pressure liquid chromatography) with cooled-autosamplers on the front end for proteomic analyses. A conventional flow UHPLC is used for metabolite analyses. These LC-MS systems represent some of the highest performing, most sensitive instruments currently available. The PMSR will continue to offer routine proteomic services such as LC-MS/MS protein identifications from gel bands or pull-down assays, and intact protein analysis by electrospray ionization. Over the last budget cycle, the PMSR has increasingly engaged in more challenging proteomics problems, particularly global quantitative comparisons of proteomes or sub-proteomes using label- free quantitation (integrated MS ion currents), quantitation using stable isotopes such as SILAC or isobaric tags (TMT or iTRAQ), and in-depth global analysis of phosphoproteome, ubiquitomes, and acetylomes. The PMSR will continue to improve its analytical capabilities for in-depth comprehensive analysis of global proteome and posttranslational modifications. The PMSR has also responded to the growing need of Cancer Center members in the field of cancer metabolism by establishing targeted relative quantitation of approximately 180 polar metabolites. To further increase its capacity in metabolomics analysis, the PMSR applied for and was recently awarded an S10 shared instrumentation grant (S10 OD023586). This grant together with Cancer Center funds were used to purchase a high-resolution Thermo Q Exactive HF-X mass spectrometer and a Vanquish UHPLC system. This instrument was installed in December 2017 and will be dedicated to untargeted (global) metabolomics, targeted metabolomics, untargeted lipidomics, targeted lipidomics and metabolic flux analysis using stable isotope tracers. The PMSR also recently purchased a Seahorse XFe96 Analyzer funded by another S10 grant (S10 OD023658). This instrument can quantify mitochondrial respiration, glycolytic activity, endogenous and exogenous fatty acid oxidation, substrate oxidation, and metabolic phenotype, providing complementary analytical capacity to the new metabolomics LC-MS/MS system. The availability of these new instruments will ensure that the PMSR remains state-of-the-art in providing proteomics, metabolomics, and lipidomics services to Cancer Center members. The major emphasis of the PMSR will be to provide expert consultation and state-of-the-art proteomics and metabolomics technologies operating at maximum performance and affordable costs to address complex cancer biology questions.